Team:Calgary/5 August 2010


Thursday August 5, 2010


OBTAINED COLONIES! Six colonies from each of the four plates were chosen to be tested for the correct construct (K135000+I13504 or K135000+I13507) by means of a colony PCR.


Today I ran a PCR of the following templates: plasmids containing MalE and MalE 31 with the signal sequence deleted. The primers are specific to signal sequence deleted MalE and MalE 31 genes.


Today, I spent the day contacting sponsors and preparing the sponsorship letters that would be used to contact the different faculties. I found colonies of the pSB1AC3 and pSB1AK3 from the registry and made overnight cultures to be Miniprepped tomorrow. Finally, I began the digest of the CpxP promoter in preparation of inserting it into a plasmid.


I did a miniprep of the plasmid switched pRFP overnights then I did a PCR of plasmid as well as a restriction digest using EcoRI and SpeI. Then I ran the 0.8% gel at 90V. The gel failed as all that was amplified was an empty plasmid. I will try to restrict again using all NEB enzymes and buffers because we ran out of invitrogen halfway. In order to get more product I am doing a gradient PCR of pRFP again and will try to PCR purify tomorrow using the vacuum method. I also designed the biography page on the wiki today.


Finished the basic preliminary structure of the wiki's front page. Will fix up some links and streamline the code tomorrow. Then it's off to start the Team page.


Today I did more research on the modelling and found out some relationships that we can use to model our IB formation. I came up with 2 ways that we can use this to create a concrete model whcih would incorporate pH and temperature, all the details in the pathways and relationships between the variables.


Today I finished off the sponsorship letters for the Kinesiology and Engineering faculties. Chris dropped these off at their respective offices on Main Campus this afternoon.

This morning I performed a restriction digest of malE31 XbaI/SpeI-cut colony 2 with PstI and EcoRI. This is to verify that malE31 was successfully biobricked, with these restriction sites. I have already successfully cut with XbaI and Pst, so we know that those ones are there. I ran the digested product on a 1% agarose gel and the results looked very good, so I sent this colony down for DNA sequencing at the facility downstairs.

Raida and I ran the gel of our PCR reaction yesterday. There was amplification of malE and malE31, but there was very limited amplification with the Ssdel primers. We determined that part of the problem may have been the use of the wrong plasmids for the reaction, so we set up a new PCR reaction using a new aliquot of the SSdel primer as well as the right plasmids. We ran a 1% agarose gel of this reactioon this afternoon. This showed some amplification in two of the lanes, but not a lot so we are going to proceed by setting up another PCR reaction with these primers in hoped of getting enough amplification to proceed with PCR Purification.